Epimedin B exhibits pigmentation by increasing tyrosinase family proteins expression,activity,and stability

Epimedin B(EB)is one of the main flavonoid ingredients present in Epimedium brevicornum Maxim.,a traditional herb widely used in China.Our previous study showed that EB was a stronger inducer of melanogenesis and an activator of tyrosinase(TYR).However,the role of EB in melanogenesis and the mechanism underlying the regulation remain unclear.Herein,as an extension to our previous investigation,we provide comprehensive evidence of EB-induced pigmentation in vivo and in vitro and elucidate the melanogenesis mechanism by assessing its effects on the TYR family of proteins(TYRs)in terms of expression,activity,and stability.The results showed that EB increased TYRs expression through microphthalmia-associated transcription factor-mediated p-Akt(referred to as protein kinase B(PKB))/glycogen synthase kinase 3β(GSK3β)/β-catenin,p-p70 S6 kinase cascades,and protein 38(p38)/mitogen-activated protein(MAP)kinase(MAPK)and extracellular regulated protein kinases(ERK)/MAPK pathways,after which EB increased the number of melanosomes and promoted their maturation for melanogenesis in melanoma cells and human primary melanocytes/skin tissues.Furthermore,EB exerted repigmentation by stimulating TYR activity in hydroquinone-and N-phenylthiourea-induced TYR inhibitive models,including melanoma cells,zebrafish,and mice.Finally,EB ameliorated monobenzone-induced depigmentation in vitro and in vivo through the enhancement of TYRs stability by inhibiting TYR misfolding,TYR-related protein 1 formation,and retention in the endoplasmic reticulum and then by downregulating the ubiquitination and proteolysis processes.These data conclude that EB can target TYRs and alter their expression,activity,and stability,thus stimulating their pigmentation function,which might provide a novel rational strategy for hypopigmentation treatment in the pharmaceutical and cosmetic industries.

Measurement and Study of Tyrosinase Activity in Potato

[Objective] The paper was to study the activity of tyrosinase,so as to provide reference for preventing browning of potatoes as well as fruits and vegetables containing tyrosinase.[Method] With potato as raw material and Na2HPO4-HCl in buffer solution（pH=7.2）as system,spectrophotometer was used to measure the absorbance of potato extract at 480nm,the curve was established,and the activity of tyrosinase was obtained.[Result] The changes of data were little under the above conditions,and the stability of tyrosinase activity was high.[Conclusion] Using spectrophotometry to determine the tyrosinase activity in potato is simple with high accuracy.

Research on Activity and Expression of Tyrosinase in Varicorhinus macrolepis

[Objective] This study aimed to analyze tyrosinase activity and its expression in Varicorhinus macrolepis. [Method] V. macrolepis was used as experimental material for the analysis and research of tyrosinase in nine kinds of organs and tissues of male and female V. macrolepis individuals by using polyacrylamide gel electrophoresis and biochemical staining method, spectrophotometry and enzyme histochemical technology. [Result] Tyrosinase exists in the liver and pancreas, intestine and spleen of female and male V. macrolepis and in the gallbladder of male V. macrolepis. Tyrosinase activities in various tissues of V. macrolepis varied largely. Specifically, tyrosinase activities in the spleen was the maximum, which was higher in female V. macrolepis than in males. According to the enzyme histochemistry results, strong positive signals of tyrosinase existed in the spleen, intestine, liver and pancreas and gallbladder of V. macrolepis, which was the strongest in the spleen. [Conclusion] In this paper, research on tissue localization of tyrosinase in V. macrolepis had been first reported, which provided theoretical basis for further exploring the functions of tyrosinase in V. macrolepis.

Study on Extraction and Catalytic Activity of Tyrosinase

Objective] Tyrosinase is a kind of polyphenol oxidase used widely, and the study of tyrosinase attracts extensive attention. [Method] ln this experiment, A conversion kinetic curve of dopa solution was obtained using absorbance of potato extract at 480 nm in Na2HPO4-HCl buffer system at a pH value of 7.2 with the vari-ation rate of absorbance against time （ΔA/Δt） as reaction rate, and the activity of tyrosinase could thus be calculated. [Result] The results showed that at the wave-length, in the Na2HPO4-HCl buffer system with a pH value at 7.2, there was little in-terference to the system, the data showed smal fluctuations, and the activity of ty-rosinase was high and stable, further providing a theoretical foundation for the de-termination and study of factors influencing tyrosinase activity. [Conclusion] The catalytic activity of tyrosinase was studied and determined with buffer solution as substrate with high accuracy and good effect.

Tyrosinase和Nestin及CD34在非典型黑素痣组织中的表达及临床意义

目的:探讨Tyrosinase、Nestin及CD34在非典型黑素痣组织中表达及意义。方法:收集2012年1月-2019年10月52例非典型黑素痣组织及20例正常皮肤组织。采用EnVision二步法检测Tyrosinase、Nestin、CD34的表达。结果:Tyrosinase和Nestin在交界痣中阳性率为100.0%和0.0%,复合痣中表皮、真皮浅层及真皮深层组织阳性率分别为100.0%,68.4%,0.0%;0.0%,10.5%,89.5%,皮内痣中真皮浅层及真皮深层组织阳性率分别为88.2%,0.0%;29.4%,88.2%,正常皮肤组织阳性率为100.0%,15.0%。非典型黑素痣细胞不表达CD34,正常皮肤组织血管内皮细胞中CD34呈强阳性。复合痣和皮内痣表皮组织痣细胞Tyrosinase阳性率显著高于真皮深层组织(P=0.0300,0.0001);复合痣和皮内痣真皮浅层痣细胞Nestin阳性率显著低于真皮深层组织(P=0.0001,0.0020)。复合痣和皮内痣的真皮浅层痣细胞中Tyrosinase和Nestin高表达率比较,差异有统计学意义(P=0.0010,0.0020);复合痣和皮内痣的真皮深层痣细胞中Nestin与Tyrosinase高表达率比较,差异有统计学意义(P=0.0000,0.0001)。结论:非典型黑素痣从表皮、真皮浅层到真皮深层Tyrosinase阳性率逐渐降低,Nestin阳性率逐渐增加,不表达CD34,Tyrosinase/Nestin的表达趋势可解释黑素细胞前体分化、迁移过程。

Down-regulation of tyrosinase,TRP-1,TRP-2 and MITF expressions by citrus press-cakes in murine B16 F10 melanoma

Objective:To investigate the suitability of citrus-press cakes,by-products of the juice industry as a source for the whitening agents for cosmetic industry.Methods:Ethylacetate extracts of citrus-press cakes(CCE)were examined for their anti-melanogenic potentials in terms of the inhibition of melanin production and mechanisim of melanogenesis by using Western Blot analysis with tyrosinese,tyrosinase-related protein-1(TRP-1),TRP2,and microphthalmia-associated transcription factor(MITF)proteins.To apply the topical agents,citrus-press cakes was investigated the safety in human skin cell line.Finally flavonoid analysis of CCE was also determined by HPLC analysis.Results:Results indicated that CCE were shown to down-regulate melanin content in a dose-dependent pattern.The CCE inhibited tyrosinase,TRP-2,and MITF expressions in a dose-dependent manner.To test the applicability of CCE to human skin,we used MTT assay to assess the cytotoxic effects of CCE on human keratinocyte HaCaT cells.The CCE exhibited low cytotoxicity at 50μg/mL.Characterization of the citrus-press cakes for flavonoid contents using HPLC showed varied quantity of rutin,narirutin,and hesperidin.Conclusions:Considering the anti-melanogenic activity and human safety,CCE is considered as a potential anti-melanogenic agent and may be effective for topical application for treating hyperpigmentation disorders.

Inhibitory Effect of Ferulic Acid on Oxidation of L-DOPA Catalyzed by Mushroom Tyrosinase

The inhibitory effect of ferulic acid on the diphenolase activity of mushroom tyrosinase and the kinetic behavior were studied with L-3,4-dihydroxyphenylalanine （L-DOPA） as substrate. The inhibitor concentration leading to 50% relative activity lost （IC50） was estimated to be 0.15 mmol·L^-1. The inhibition mechanism obtained from Lineweaver-Burk plots shows that ferulic acid is a competitive inhibitor and the inhibition of tyrosinase by ferulic acid is a reversible reaction. The equilibrium constant for ferulic acid binding with the tyrosinase was determined to be 0.25 mmol·L^-1 for diphenolase. Keywords tyrosinase, ferulic acid, kinetics, inhibition, L-DOPA, diphenolase

Fluorescence quenching method for the determination of catechol with gold nanoparticles and tyrosinase hybrid system

The determination method of catechol by fluorescence quenching was developed.The assay was based on the combination of the unique property of gold nanoparticles with tyrosinase enzymatic reaction.In the presence of tyrosinase,the fluorescence of gold nanoparticles was quenched by catechol which can be employed to detect catechol.Under the optimal conditions,a linear range 5.0×10^(-7)-1.0×10^(-3) mol L^(-1) and a detection limit 1.0×10^(-7) mol L^(-1) of catechol were obtained.o-Quinone intermediate produced...

S-100、HMB-45和Tyrosinase在恶性黑色素瘤中的表达及其意义

目的探讨S-100、HMB-45和Tyrosinase在恶性黑色素瘤(MM)诊断和鉴别诊断中的应用价值。方法采用免疫组化检测S-100、HMB-45和Tyrosinase在28例MM、17例良性痣中的表达,比较三种抗体对MM检测的敏感性和特异性。结果 S-100表达的敏感性较高(96.43%),但特异性低(5.89%);HMB-45表达的特异性较高(76.47%);Tyrosinase表达的敏感性较高(96.43%)。结论选择Tyrosinase和HMB-45联合标记,可有效提高恶性黑色素瘤诊断的敏感性和特异性。

Phenol content,antioxidant and tyrosinase inhibitory activity of mangrove plants in Micronesia

Objective:To find out and compare the in vitro antioxidant and tyrosinase inhibitory activities of two species of mangrove plants.Methods:Mangrove samples were harvested at the shoreline on the island of Weno,Chuuk State in Micronesia.The phenol content,antioxidant activity(based on DPPH-free radical scavenging)and tyrosinase inhibitory activity in different tissues(leaves,barks and roots)of Rhizophora stylosa(R.stylosa)and Sonneratia alba(S.alba),collected from the island of Weno.Results:Total phenol content ranged from 4.87 to 11.96 mg per g of freeze dried samples.The highest antioxidant activity was observed in R.styiosa bark(85.5%).The highest tyrosinase inhibitory activity was found in S.alba bark.Also,total phenol content and antioxidant activity were higher in methanol extracts than in aqueous extracts.Conclusions:Taken together,the results of this study proved that mangroves can be excellent sources of antioxidant compounds.

Attachment of tyrosinase on mixed self-assembled monolayers for the construction of electrochemical biosensor

A mixed self-assembled monolayers （SAMs） of thioctic acid （T-COOH） and thioctic acid amide （T-NH2） were used to immobilize tyrosinase for fabricating biosensor. The results showed that the mixed SAMs prepared from solution at the ratio of 1：4 provided an excellent microenvironment for enzymatic reaction between tyrosinase and substrate. The biosensor exhibited a fast response and high sensitivity for sensing substrate.

Phenolic Constituents with Antioxidative,Tyrosinase Inhibitory and Anti‑aging Activities from Dendrobium loddigesii Rolfe

Aqueous ethanol extracts of powdered stems of Dendrobium loddigesii afforded three new phenolics including threo-7-Oethyl-9-O-(4-hydroxyphenyl)propionyl-guaiacylglycerol(1),(R)-4,5,4ʹ-trihydroxy-3,3ʹ,α-trimethoxybibenzyl(2)and(S)-5,5′,7-trihydroxy-3′,4′-dimethoxyflavanone(3),together with eleven known analogues.Their structures were determined by extensive spectroscopic analysis.To identify natural antioxidants,whitening,and anti-aging agents,the abilities of these phenolics were assessed to scavenge the 1,2-diphenyl-2-picrylhydrazyl(DPPH)radical,their abilities to inhibit tyrosinase production,and their abilities to stimulate collagen production by human dermal fibroblasts-adult(HDFa)assay.It was found that compounds 1,4-8,13 and 14 exhibited significant DPPH radical scavenging activities,compound 10 exhibited tyrosinase inhibitory activity(IC_(50)37.904μg/mL),and compound 9 showed significant collagen production with an EC_(50)value of 3.182μg/mL.These results suggest that phenolic constituents from D.loddigesii may be candidate antioxidants,skin-whitening and/or anti-aging agents.

Catalysis-based specific detection and inhibition of tyrosinase and their application

Tyrosinase is an important enzyme in controlling the formation of melanin in melanosome,and plays a key role in the pigmentation of hair and skin.The abnormal expression or activation of tyrosinase is associated with several diseases such as albinism,vitiligo,melanoma and Parkinson disease.Excessive deposition of melanin could cause diseases such as freckles and brown spots in the human body,and it is also closely related to browning of fruits and vegetables and insect molting.Detecting and inhibiting the activity of tyrosinase is of extraordinary value in the progress of diagnosis and treatment of these diseases.Therefore,many selective optical detection probes and small molecular inhibitors have been developed,and have made significant contributions to the basic and clinical research on these diseases.In this paper,the detection and inhibition of tyrosinase and their application in whitening products are reviewed,with special emphasis on development of fluorescent probes and inhibitors.Hopefully,this review will help design more efficient and sensitive tyrosinase probes and inhibitors,as well as shed light on novel treatment of diseases such as melanoma.

Isolation, Identification and Tyrosinase Inhibitory Activities of the Extractives from Allamanda cathartica

Tyrosinase inhibitory activity of the extractives from A. cathartica was examined and their new bioactivity and potent active compounds were identified. Five compounds, glabridin, new lignan, kaempferol, naringenin, and allamandicin, were isolated by a series of chromatography, and identified by NMR and LC-MS. Among them, glabridin had the high-est tyrosinase inhibitory activity (IC50:2.93 μM) which is 15 times stronger than that of kojic acid used as positive con-trol (IC50:43.7 μM). Moreover the lignan was indentified as 1-[3-(4-allyl-2,6-dimethoxyphenoxy)-4-methoxyphenyl] propane-1,2,diol which was a novel lignan.

Mushroom Tyrosinase Inhibition and Antimelanogenesis Activities of <i>Bacopa monnieri</i>(L.) Methanol Extract in B16F10 Melanoma Cells

Melanocytes that form stratum basale of skin epidermis express tyrosinase enzyme, which catalyzes initial two rate-limiting steps in the biotransformation of tyrosine into dark pigment called melanin. Even today, Tyrosinase inhibitors are among the promising candidates in cosmetic industry for skin-lightening formulations. Overexpression of tyrosinase causes excess melanin biosynthesis and deposition resulting in dark skin color. Moreover, localized overexpression of tyrosinase cause variety of hyperpigmentation disorders like melanoma, melasma, chloasma, dark patches, liver patches, etc. There has been a renewed interest in the natural products as main ingredients in the formulation of safe products for skin-whitening and treatment options for hyperpigmentation disorders. In the present communication, the results of our investigations on tyrosinase inhibition, modulation of intracellular tyrosinase and melanin levels in cultured B16F10 melanoma cells by Bacopa monnieri (L.) methanol extract (BME) are presented and discussed as safe option for skin lightening and to treat hyperpigmentation disorders. BME showed 11%, 29%, 54% and 80% inhibition of mushroom tyrosinase activity at an initial 100, 200, 400 and 600 μg of extract. Treatment of α-melanocyte stimulating hormone (α-MSH) stimulated cultured murine melanoma B16F10 cells with 100 μg/ml of the extract showed a decrease in the levels of cellular melanin and cellular tyrosinase content by 22% and 46% respectively. The cytotoxicity studies by MTT assay revealed that the LC50 of the BME is ≥1000 μg/ml in cultured mouse melanoma B16F10 and HEK293 cells.

The tyrosinase gene family and albinism in fish

Tyrosinase exists universally in organisms and is a characteristic enzyme of melanocytes. Tyrosinase family genes in vertebrates consist of 3 related members; tyrosinase （TYR, Tyr）, tyro sinase-related protein-1 （TRP-1, Tyrp 1）, and tyro sinase-related protein-2 （TRP-2, Tyrp2, Dct）. These proteins catalyze melanin biosynthesis in pigment cells and play important roles in determining vertebrate coloration. Transcription of the TYR and TRP genes is useful for studying neural crest and optic vesicle cell migration and differentiation during emblyogenesis and important in pigment rescue in fish. In this paper, the structure of gene and protein molecular evolution, function and roles of the TYR family in fish were reviewed.

Studying on Tyrosinase Inhibition Activity of Some Vietnamese Folk Plants Aims to Use in Skin-Whitening Cosmetics

White mulberry Morus alba and jackfruit Artocarpus heterophyllus are two of seven chosen Vietnamese folk plants which showed the highest tyrosinase inhibition activity. Besides, both of them showed antioxidant and antibacterial activities. Especially, the Sulforhodamine B (SRB) assay test result of the root core extract of M. alba figured out its non-toxicity on foreskin fibroblasts.

Kinetic Characterization of Tyrosinase-catalyzed Oxidation of Four Polyphenols

Phenolic compounds such as chlorogenic acid,cryptochlorogenic acid,neochlorogenic acid and caffeic acid are widely distributed in fruits,vegetables and traditional Chinese medicines with a wide range of biological activities.Tyrosinase plays a critical role in the food industry,but recent studies have proposed unexplored aspects of clinical application.Tyrosinase-catalyzed oxidation of four polyphenols as well as its underlying mechanism remains unclear.In the current work,we investigated the kinetic properties of tyrosinase-catalyzed oxidation of the four polyphenols of interest.To measure the unstable o-quinone products,an analytical method using 3-methyl-2-benzothiazolinone hydrazone(MBTH)was established.The optimal incubation time,buffer pH,temperature and enzyme concentration for the enzyme activity in the presence of each polyphenol of interest were investigated.Under the final optimized conditions,the kinetics and substrate specificity of four polyphenols were examined.K inetic data showed that tyrosinase had the greatest substrate affinity to chlorogenic acid compared with its isomers and caffeic acid.The catalytic efficiency with chlorogenic acid was 8-to 15-fold higher than that with the other 3 polyphenols.Molecular docking study demonstrated that the tight binding of chlorogenic acid at the peripheral site should be the major reason for the specificity to chlorogenic acid.In light of this,the rational design of high-affinity inhibitors against tyrosinase may focus on the binding of both the Cu site and peripheral site.This study will supply a basis for the selection of phenolic acids in food industry and health carc.

Development and Application of a Tyrosinase-Based Time-Temperature Indicator(TTI) for Determining the Quality of Turbot Sashimi

Time-temperature indicators(TTIs) are convenient intuitive devices that are widely used to predict food quality. The aim of this study is to develop a new simple device which can be attached to food packages as a quality indicator for turbot sashimi. In this study, a solid TTI based on the reaction between tyrosinase and tyrosine was developed. The Arrhenius behavior of this enzymatic TTI was studied. The kinetics of the tyrosinase-based TTI was investigated in the form of color change from colorless to dark black induced by the enzymatic reaction. The mathematical formula for the color alterations as a function of time and temperature was established. The longest indication time for the developed TTI was 50 hours at 4℃. The activation energy of the tyrosinase-based TTI was 0.409 k J mol^(-1). The suitability of the tyrosinase-based TTI was validated for turbot sashimi using total plate count. The feasibility of using this TTI as a quality indicator for turbot sashimi was assessed based on the activation energy and indication time. Therefore, the tyrosinasebased TTI system developed in this study could be used as an effective tool for monitoring the quality changes of turbot sashimi during the distribution and storage.

Tyrosinase,a new innate humoral immune parameter in large yellow croaker(Pseudosciaena crocea R)

We evaluated the immune response to infection with a pathogen in large yellow croaker(Pseudosciaena crocea Richardson).The fish were given an intraperitoneal(i.p.) injection of Vibrio parahaemolyticus or sterile sea water(control).We collected blood sera from the fish 0.17,1,2,4,8,12,or 16 d after injection(dpi).We measured tyrosinase activity and the concentrations of lysozyme,NOS,and antibodies.Serum tyrosinase activity was significantly higher at 0.17 and 4 dpi than in the control group,and peaked at 8 dpi.Lysozyme activity was significantly higher at 2 and 12 dpi than in the control group,but lower at 16 dpi.There is no statistical difference in the level of nitric oxides synthase(NOS) activity or antibodies between the control and injection groups.This is the first report of the tyrosinase activity in the serum of large yellow croaker.Our results indicate that tyrosinase plays an important role in the immediate immune defense against V.parahaemolyticus in large yellow croaker.Tyrosinase is a candidate parameter for investigation of fish innate immune defense.